tretinoin and Graves-Disease

It is well known that regulatory T cells (Tregs) are abnormal in Graves' disease (GD) and play crucial roles in the breakdown of immune tolerance and GD development. However, there are controversies about whether the quantity and/or function of Tregs is aberrant in GD. The molecular mechanism of Tregs abnormality and its effects on GD development was still unclear, until now.. MiRNAs play important roles in the function and development of the immune system including Tregs. To reveal the Tregs abnormality and its molecular mechanism in GD, we systematically studied the quantity and immunosuppressive function as well as the differential expression profiles of miRNA and mRNA of Tregs in newly diagnosed patients with GD using TaqMan miRNA array and mRNA microarray.. Our results showed that the quantity and immunosuppressive function of Tregs in initial patients with GD was significantly decreased. More importantly, the retinoic acid (RA) pathway was markedly suppressed and its agonist, all-trans retinoic acid, could notably improve the quantity and immunosuppressive function of Tregs from patients with GD in vitro. In addition, many other pathways including protein ubiquitination and circadian rhythm were also significantly regulated in Tregs of GD.. This integrative study first revealed the expression profiles of mRNA/miRNA in Tregs of initial GD and RA pathway might play important roles in GD development. Our results implied that all-trans RA, which had been used for a long time in the clinical setting, had potential value in the treatment of GD and was worthy of additional study.

Topics: Adult; Female; Graves Disease; Humans; Male; Microarray Analysis; MicroRNAs; Middle Aged; RNA, Messenger; Signal Transduction; T-Lymphocytes, Regulatory; Tretinoin

In this study, we evaluated by reverse transcription-polymerase chain reaction (RT-PCR) the expression pattern of retinoic acid receptors (RAR) alpha, beta, and gamma and cellular retinoic acid binding protein-I (CRBP-I) genes in 12 primary cultures of fibroblasts (F) from orbital tissue of Graves' ophthalmopathy (GO) patients. We also studied the in vitro effects of all-trans retinoic acid (RA) and N-(4-hydroxyphenil)-retinamide (4HPR), a less toxic and better tolerated synthetic derivative of RA, on cell morphology, growth, apoptosis, and cyclic adenosine monophosphate (cAMP) accumulation. All primary cultures expressed RAR alpha, beta, gamma, and CRBP-I. FGO treated with RA and 4HPR (10(-7) mol/L) presented morphologic changes and significantly inhibited cell growth after 72 hours. At 96 hours of drug exposure, apoptosis was detected in 15% and 50% of RA- and 4HPR (10(-7) mol/L)-treated cells, and p53 protein increased in cell lysates. 4HPR induced a 70% decrease of Bcl-2 protein. After 30 minutes of RA and 4HPR (10(-7) mol/L) exposure, a 20% decrease of basal cAMP accumulation was seen, and forskolin cAMP-induced increase was abolished. The expression of RAR alpha, beta, gamma, and CRBP-I in primary cultures of FGO indicates that they are targets for retinoids. Moreover, we show that RA and 4HPR are able to induce morphologic changes, inhibition of cell growth, and apoptosis in FGO exerting their effects through RAR-modulated pathways. The rapid inhibition of cAMP accumulation indicates that a novel nonclassic retinoid pathway may also be involved. Finally, the potent in vitro effects of 4HPR, a retinoid derivative with fewer adverse reactions in vivo, could justify further investigations on a clinical application of retinoids in GO.

Topics: Adult; Aged; Apoptosis; Blotting, Western; Cell Division; Cells, Cultured; Cyclic AMP; Electrophoresis, Polyacrylamide Gel; Female; Fenretinide; Fibroblasts; Genes, bcl-2; Graves Disease; Humans; Male; Middle Aged; Orbit; Receptors, Retinoic Acid; Retinol-Binding Proteins; Retinol-Binding Proteins, Cellular; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transcription, Genetic; Tretinoin; Tumor Suppressor Protein p53

We have investigated the in vitro expression of membrane and soluble intercellular adhesion molecule-1 (ICAM-1) by human thyroid cells from 20 patients with Graves' disease and 5 normal subjects. Membrane ICAM-1 was not detected by flow cytometry analysis in non-cultured thyrocytes from either normal or Graves' disease tissues. It appeared on thyroid cells after a 24-h culture in monolayers and showed a regular dose-dependent increase. The same results were obtained with soluble ICAM-1 (sICAM-1) in culture media from cells cultured in monolayers, vesicles or follicles. No change was obtained with different concentrations of fetal calf serum added to the media. Coculture of Graves' disease thyrocytes with autologous peripheral blood lymphocytes (PBL) or intrathyroidal lymphocytes (ITL) enhanced the expression of both membrane and sICAM-1 whatever the culture model. When normal thyrocytes were cocultured with PBL, sICAM-1 increased but with ITL sICAM-1 remained unchanged. High concentrations of gamma interferon induced an increase of both membrane and sICAM-1 in the three culture models. However the increases were greater with vesicles and follicles. Only sICAM-1 levels were raised with 0.1, 1 and 10 microM retinoic acid. These results suggest that ICAM-1 appears in culture, possibly due to mechanical effects such as adherence to plates and cell-to-cell contacts. Moreover, its expression is modulated by several factors such as cytokines or retinoic acid. Further investigations are needed to establish whether ICAM-1 is really involved in the pathogenesis of Graves' disease.

Topics: Adult; Blood; Cell Culture Techniques; Cell Membrane; Cell Separation; Cell-Free System; Cells, Cultured; Coculture Techniques; Female; Graves Disease; Humans; Intercellular Adhesion Molecule-1; Interferon-gamma; Lymphocyte Activation; Lymphocyte Subsets; Male; Middle Aged; Reproducibility of Results; Solubility; Thyroid Gland; Tretinoin

The effect of retinoic acid (RA) on thyroid peroxidase (TPO) and thyroglobulin (Tg) gene expression was investigated in cultured human thyrocytes. Thyrocytes dispersed from Graves' thyroid tissues were incubated with TSH 5mU/ml and RA 0, 0.01, 0.1, 1.0 microM for 72 h respectively. The samples were then subjected to Northern gel analysis. Northern gel analysis using the specific cDNA probes showed that RA suppressed the accumulation of TPO and Tg mRNA stimulated by TSH in a time- and dose-responsive manner. Furthermore, RA inhibited forskolin and 8-Bromo-cyclic-AMP-induced TPO and Tg gene expression, suggesting a distal action site for these cAMP mediated gene expressions. Immunoprecipitation analysis using the specific monoclonal antibodies showed that TSH increased newly synthesized 100, 75, 36-kDa [35S] TPO. The increased de novo TPO was markedly inhibited by RA. Tg secretion from monolayer cultures was measured by radioimmunoassay. RA also inhibited TSH-induced Tg secretion in a dose dependent manner. RA did not affect [3H] thymidine uptake into primary cultured human thyrocytes. In conclusion, RA inhibits the synthesis of TPO and Tg via the suppression of thyroid-specific gene expression although the exact site of RA action on these genes in human thyroids remains to be further elucidated. These results suggest that RA may play a regulatory role in Tg and TPO gene expression, subsequently resulting in the suppression of thyroid hormone synthesis.

Topics: 8-Bromo Cyclic Adenosine Monophosphate; Blotting, Northern; Cells, Cultured; Colforsin; DNA; DNA Probes; Gene Expression; Graves Disease; Humans; Immunosorbent Techniques; Iodide Peroxidase; RNA, Messenger; Thyroglobulin; Thyroid Gland; Thyrotropin; Tretinoin